鱼肉DNA提取方法比较及物种鉴定研究

Comparison of DNA Extraction Methods and Species Identification of Fish Meat

摘要:

本研究采用改进煮沸法（BP法）以及传统蛋白酶K法（TP法），从16种鱼肉样品中提取DNA，分析比对两种方法提取DNA的浓度、纯度、以及普通PCR和荧光PCR的效果。同时，运用BP法从70个市售鱼肉产品中提取总DNA，并采用DNA条形码技术进行物种鉴定。结果表明，BP法提取DNA耗时较短，对于大部分鱼肉样品，BP法提取的DNA浓度低于TP法，但两种方法提取的DNA A260/A280无显著差异，均满足普通PCR和荧光PCR扩增要求。另外，BP法提取的DNA可用于市售鱼肉产品DNA条形码鉴定，通过比对发现，81.43%的鱼肉产品可匹配到相似度≥98%的物种，在种水平和属水平的鉴定成功率分别为48.57%和72.86%。BP法在提取DNA过程中可缩短时间，降低提取成本，便于大批量样品的检测。

Abstract:

In this study, an improved boiling method (BP method) and traditional proteinase K method (TP method) were used to extract DNAs from 16 kinds of fish meat samples. Then, the concentration and purity of the extracted DNAs as well as the performance of the common polymerase chain reaction (PCR) and fluorescence PCR were compared. In the mean time, the total DNA was extracted from 70 commercially available fish products using the BP method and analyzed for species identification using DNA barcode technology. The results showed that the extraction of DNA by the BP method required a shorter time. For most fish samples, the concentration of the DNA extracted by the BP method was lower than that by the TP method, but the two methods led to insignificant difference in the A260/A280 ratio of the extracted DNAs, thus, met the requirements for common and fluorescent PCR amplification. Moreover, the DNA extracted by the BP method could be used for DNA barcode identification of commercially available fish products. By comparison, 81.43% of the fish products could be matched with a similarity degree ≥ 98%, and the identification success rates at the species and genus levels were 48.57% and 72.86%, respectively. The BP method can shorten the DNA extraction time, reduce the extraction cost, and facilitate the detection of large quantities of samples.